User:Cynthia Pruss/Sandbox 1

=Von Willebrand Factor A2 Domain= A2 domain A2 with Y1605/M1606 VWF73 VWF115 Amino Acids of Interest Y1584 G1643



This is the crystal structure of the VWF A2 domain, PDB ID 3GXB, elucidated by Zhang, et al. Von Willebrand factor (VWF) is a large multimeric glycoprotein with a critical role in maintaining hemostasis. VWF serves as the carrier protein for the plasma coagulation protein factor VIII (FVIII) and promotes platelet adhesion and aggregation at the sites of vascular damage. The hemostatic potential of VWF greatly increases with the multimer size, which is tightly regulated in vivo by the metalloprotease ADAMTS13.

ADAMTS13 is the thirteenth member of the A disintegrin-like and metalloprotease with thrombospondin type 1 motif family of proteases. ADAMTS13 cleaves VWF between Tyr1605-Met1606, and maintains plasma VWF within a normal molecular weight range by cleaving high molecular weight (HMW) multimers soon after they are secreted into the plasma, preventing spontaneous platelet agglutination.

The A2 domain is not stabilized by disulfide bonds within VWF, unlike A1 and A3. In addition, the A2 domain lacks the highly conserved α4 helix, which has been replaced with the flexible and dynamic α4-less loop. This leads to low resistance to unfolding, giving the A2 domain the function of a shear bolt. A shear bolt breaks above a designed force threshold, to protect other parts of a machine from accidental damage. Similarly, the A2 domain unfolds when present in VWF multimers that experience high tensile force and is cleaved by ADAMTS13, resulting in down regulation of the hemostatic activity. VWF will only be exposed to peak shear forces intermittently in the arterioles in normal circulation, but when tethered, VWF will undergo longer, higher shear rates, causing the A2 domain to unfold and allowing ADAMTS13 access to the A2 domain’s central cleavage site. These shear forces will increase at sites of vascular damage with a growing thrombus that narrows vessel diameter. ADAMTS13 serves to halt the growth of the thrombus to prevent vessel occlusion, as has been demonstrated in vitro. The minimal substrate for ADAMTS13 cleavage is the A2 domain’s D1596-R1668, VWF73. Several exosites in ADAMTS13 interact with the VWF A2 domain to contribute to substrate specificity and enhance cleavage efficiency. The TSP1 spacer domain in ADAMTS13 interacts with VWF residues between Gln1624 and Arg1668. The exosite binding was further refined to identify three additional exosites, the first involving VWF1596-1623, the second VWF1642-1652, and the third VWF1653-1668, that are exposed when the VWF A2 domain is unfolded.

Type 2A group II von Willebrand disease mutations have increased susceptability to ADAMTS13-mediated proteolysis. These are located in the A2 domain. These include R1597W.

Y1584C is a type 1 von Willebrand disease mutation that has also demonstrated increased ADAMTS13-mediated proteolysis. Atoms within 5 angstroms are highlighted. The red circles represent the oxygens in free water molecules around Y1584. The A2 domain has several naturally occuring single nucleotide polymorphisms that alter the amino acid sequence, and also alter ADAMTS13 mediated cleavage. These include Q/H1571, P/T1601, and G/S1643.

Amino Acids of Interest